DESCRIPTION ABOUT THE MOLECULAR PATHWAY OF CANCER, DIFFERENT MOLECULAR MARKERS, METHOD OF DETECTION AND ITS ROLE IN MANAGEMENT OF NSCLC

1. BY Dr DEEPAK KUMAR DAS
MOD- PROF. S. GHOSHAL
DEPT. OF RADIATION ONCOLOGY
PGIMER, CHANDIGARH

2. MOLECULAR BASIS OF CANCER

3. CLINICAL USEFULLNESS OF MOLECULAR
MARKERS
 Diagnosis of tumour susceptibility
 Diagnosis of tumour micrometastasis, including early diagnosis and
molecular staging
 Evaluation of tumour profiling, such as tumour malignancy and drug
sensitivity
 Molecular targeted therapy

4. DIAGNOSIS OF TUMOUR SUSCEPTIBILITY
 Single nucleotide polymorphism(SNP) occurs once per 1000
nucleotide in an individual.
 These polymorphisms occur within enzymes resulting in change
of activity.
 The metabolic activation of carcinogens by CYP1A1, CYP2A6 and
CYP2E1, and their detoxification and inactivation by GSTM1 is
altered due their genetic polymorphism.

5. DIAGNOSIS OF TUMOUR MICROMETASTASIS
 The presence of tumour micrometastasis is helpful in early
diagnosis and molecular staging of tumour.
 Lung cancer is diagnosed as tumour micrometastasis when gene
alteration such as p53, Kras and p16 mutation, and telomerase
activity are identified from sputum, blood and bronchoalveolar
lavage fluid.
EVALUATION OF TUMOUR PROFILING
 There is no novel prognostic factor with a ultility of
determining the prognosis of lung cancer patients
equal to staging.
 But it is possible to estimate the individual drug susceptibility
( tumour drug susceptibility)

6.  The destruction of cell by cisplatin requires binding of the drug to DNA and
formation of platinum-DNA adducts.
 These adducts produce cross link between DNA strands and prevents replication.
 Nucleotide excision repair has central role in DNA repair and is associated with
resistance to platinum based chemotherapy.
 The excion repair cross complementation group 1(ERCC 1) plays an important role
in nucleotide excision repair pathway that identifies and removes cisplatin induced
DNA adducts.

7. PREDICTIVE ROLE OF THYMIDYLATE SYNTHASE
 Thymidylate synthase, an important enzyme in DNA biosynthesis is
one of the target enzyme for antifolate drugs.
 High levels of TS has been shown to be higher in squamous cell
carcinoma and correlate with resistance to antifolate drugs such as
premetrexed
 Premetrexed along with cisplatin is approved as first line therapy in
advanced lung cancer patients with nonsquamous histology.

8. TARGETED THERAPY
 Drugs targeted at pathways, processes and physiology which are
uniquely disrupted in cancer cells:
 Receptors
 Genes
 Angiogenesis
 Tumour PH
 In reality these pathways are not so distinct

9. IDEAL MOLECULE FOR TARGETED THERAPY
 The molecule is uniquely expressed in cancer cells; hence the
therapeutic agent will specifically target the cancer and not the normal
cells.
 The molecule is important for the maintenance of the malignant
phenotype; therefore once the targeted molecule has been effectively
disabled, the cancer cell will not be able to develop resistance against
therapeutic agent by suppressing its function or expelling the targeted
molecule from the cell.
The degree to which target molecules do not embody these characteristics
coupled with nonspecificity of the therapeutic agent determines, in part,
the limitations of current targets and agents.

10. CLASSIFICATION OF MOLECULAR TARGETED THERAPY
I-Function directed therapy
 Cell signaling targeted therapy
1-Blocking of ligand receptor binding
a- EGFR family
-EGFR1 targeted therapy
- Her-2-neu targeted therapy
b- Vascular endothelial growth factor(VEGF)
c- IGF1R
2- Inhibition of RTKs
- Erlotinib
- Geftinib
3- Inhibition of intracellular signaling proteins and protein kinases
a- Bcr-Abl tyrosine kinase- Imatinib, dasatinib
b- Raf/MAP kinase pathway- sorafenib
c- PI3K, AKT, and mTOR pathway inhibitors- Everolimus

11. Angiogenesis targeted therapy
 Protein degradation targeted therapy- Bortezomib
 Immune modulation targeted therapy
II- Phenotype targeted therapy
1-Nonreceptor protein directed MoAbs
a- Unconjugated antibodies- rituximab
b- Conjugated antibodies
2- Immunotoxins
3- Cancer vaccines

12. MOLECULAR MARKERS

16. EGFR DETECTION METHODS

18.  VeriStat is a commercially available serum or plasma based test utilizing matrix
assisted laser desorption ionisation(MALDI) mass spectroscopy methods.

19.  VeriStat assigns each spectrum a binary classification of good or poor.
 Patients classified as good has a better outcome with the EGFR TKI therapy than
clasdsified as poor.
 A predictive marker

20. EGFR PATHWAY

21. TARGETING EGFR
 By monoclonal antibodies that target extracellular domain of the
receptor or by molecules that bind ATP site and inhibit tyrosine kinase
activity of the receptor
 About 10% of patients with NSCLC have activating mutation in ATP
binding pocket of EGFR which give rise to prolonged activity in
response to ligands such as EGF or TGF-beta
 Activating mutations are primarily seen in patients having minimal or
no smoking history, Asian women and more common in
adenocarcinoma than squamous cell carcinoma
 Patients with wild type, overexpressed EGFR are more likely to show a
response of growth restraint but not regression

22. FREQUENCY OF MUTATIONS

23. IPASS: STUDY DESIGN
Eligibility criteria
- Age 18 years or older
-Stage IIIB or IV disease
- Adenocarcinoma histology
- Non smokers or light smokers
- No previous chemo or biologic
Or chemotherapy
-
1:1 randomisation
Geftinib 250 mg /day or
Pacli(200 mg/m2)plus carbo 5 or 6 AUC
Primary end point
- PFS

24. RESULTS
GEFTINIB PACLITAXEL PLUS
CARBOPLATIN
12 Month rate of PFS 24.9% 6.7%
Adverse reaction -Rash or acne(66.2%)
-Diarrhoea(46.6%)
-Neurotoxic effects(69.9%)
-Neutropenia(67.1%)
-Alopecia(58.4%)

26. NEJ002
No significant difference in overall survival is observed betweengeftinib and
Paclitaxel/carboplatin probably due to high rate of crossover use of geftinib in the
Pacli/carbo group

27. INTEREST TRIAL
 Locally advanced or metastatic NSCLC previously treated with
platinum based chemotherapy
Docetaxel 75
mg/m2 every 3
weely
Geftinib 250
mg/day
MEDIAN SURVIVAL 8 month 7.6 month
MEDIAN SURVIVAL
( in patients with
EGFR mutation)
7.5 month 8.4 month

28. INCLUSION CRITERIA
 Age equal to or more than 18 years
 Stage IIIB or IV NSCLC
 Confirmed activating mutation of EGFR

29. Erlotinib(150
mg/day) until
disease progression
or toxic side effects
4 cycles of gemcitabine
and carboplatin
Median PFS 13.1 month 4.6 month
Grade 3 or 4 side effects Increased ALT
Skin rash
Neutropenia
Thrombocytopenia

30. RESISTANCE TO TKIs
SECONDARY MUTATION TO EGFR
 Most common of these mutations is T790M
 Threonine at position 790, commonly referred as “gate keeper”residue
owing to its central location within ATP binding pocket is commonly
altered in NSCLC.
 Mutation of this residue to methionine(T790M) prevents incorporation
of both geftinib and erotinib due to steric constraints produced due to
decreased pocket size

31. ACTIVATION OF COMPENSATORY ONCOGENES
 Amplification and/or mutation of oncogenes with redudant function
to EGFR have emerged as another key mode of resistance to TKIs.
The best characteristic mechanism of this type is amplification of MET.

32. EPIGENETIC IDENTITY SWITCHING
 In presence of prolonged exposure to TKIs, cells undergo multistep
process of nonrandom epigenetic switching to an alternative cellular
identity.
 By switching to alternate TKs or reversion to cancer stem cell like
identity

33. IRREVERSIBLE TKIs
 These form irreversible, covalent attachments to the EGFR kinase domain.
 Due to covalent binding, they may have more activity against T790M or
secondary mutations against which first generation TKIs are ineffective.

34. AFATINIB
 Most extensively studied of 2nd generation TKIs
 Active against both wild type and mutant forms of EGFR
 Active against T790M
 Dose- 50 mg orally daily
 Dose limiting side effects include rash and dyspnea secondary to
pneumonitis

35. LUX-Lung 1

36. Afatinib Placebo
PFS 3.3 month 1.1 month
Partial response 7% 0.5%
Stable disease 51% 18%
Disease control
rate
58% 19%
RESULTS

37. LUX-Lung 2

38. All patients
N=129
EGFR mutation
DEL 19/L585R
Overall response rate 61% 65%
Disease control rate 86% 88%
Median PFS 14 month 15 month
Median OS 24 month _
RESULTS

39. LUX-Lung3
Afatinb 40 mg/day v/s 6 cycle of premetrexed plus cisplatin

40. (FOR ALL PATIENTS)
FOR PATIENTS WITH COMMON
MUTATION(del19/L858R)

41. LUX-Lung 4
 Of 62 patients, 51 patients had acquired resistance to Geftinib /or Erlotinib
 Only 5 had confirmed objective response.
 Median PFS is 4.4 month
 Median OS is 19 month

42. LUX-Lung 5
Patients with advanced
NSCLC whose disease
progressed after 1st line
chemotherapy and an
EGFR TKIs: either
erlotinib or geftinib( after
12 weeks of treatment)
Afatinib
Progression after
12 weeks of
afatinib
monotherapy
Afatinib plus
chemotherapy
Afatinib

43. (RESULT)

44. LUX-Lung6

46. (cough)
(dyspnea)

47. LUX-Lung 7
INCLUSION CRITERIA
•Male and female patient aged 18 to 90 years
• Stage IIIB/IV adenocarcinoma of lung
•Documented EGFR mutation DEL19 and/or L585R
•At least one measurable lesion according to RACIST 1.1
•ECOG performance status 0 or 1

48. LUX-Lung 8

50. OTHER 2nd GENERATION TKIs
NERATINIB
 An oral irreversible inhibitor of both EGFR and HER2
 Active against EGFR T790M
 Maximum tolerated dose is 320 mg daily
DACOMITINIB
An irreversible pan-HER tyrosine kinase inhibitor with activity against EGFR
 Maximum tolerated dose is 45 mg orally daily

51. ALK PATHWAY

52. REARRANGEMENT OF ALK
 Found in 3-5% of NSCLC
 Also found in anaplastic large cell lymphoma, neuroblastoma, IMT
 Found in younger patients
 More prevalent in patients having no history of smoking or light
smoking
 Adenocarcinoma histology
FUSION PARTNERS

53. PROFILE 1014
Crizotinib is an oral small molecule tyrosine kinase inhibitor of ALK, MET and
ROS1 kinases.

54. INCLUSION CRITERIA
ALK positive
 Advanced NSCLC
 No previous systemic treatment
CRIZOTINIB(250
mg twice daily)
Premetrexed 500
mg/m2 plus
cisplatin 75 mg/m2
or carboplatin AUC
5 or 6 every 3
weekly upto 6 cycle
PFS 10.9 Month 7 month
Objective response
rate
74% 45%
Most common
adverse event
-Vision disorder
- diarrhoea
- nausea
- edema
-Nausea
- fatigue
-Vomiting
-Decreased appetite
RESULTS

56. REARRANGEMENT OF ROS-1
 ROS-1 is activated by chromosomal rearrangement found in variety of
human cancer like NSCLC, cholangiocarcinoma, gastric cancer, ovarian
cancer, and GBM.
 Rearrangement leads to fusion of portion of ROS-1 that includes
tyrosine kinase domain with 1 of 12 partner proteins.
 The resulting fusion kinases are constitutively activated and leads to
cellular transformation.
 Found in 1% of NSCLC.
 Found in patients who have never smoked or history of light smoking.
 Adenocarcinoma histology

58.  N= 50
Advanced NSCLC
 Positive for ROS-1
RESULTS
Objective response rate was 72%
 Median duration of response was 17.6 months
Median PFS was 19.2 months

59. BEVACIZUMAB
 Angiogenesis is one of the hallmark of cancer.
 Vascular endothelial growth factor(VEGF) is the major regulator of
angiogenesis.
 Increased expression of VEGF is found in most of human tumors
including non small cell cancer.
 Humanised variant of monoclonal antibody against VEGF
(bevacizumab) has shown clinical activity against human cancer.

60. PC Bevacizumab plus
PC
Median survival 10.3 month 12.3 month
Median PFS 4.5 month 6.2 month
There were 15 treatment related death in chemo plus bevacizumab arm including
5 from pulmonary haemorrhage.

61. KRAS PATHWAY

62. SELUMETINIB
 KRAS mutation occurs in 20-30% of NSCLC patients.
 Selumetinib blocks the MAPK kinase(MEK) which is part of KRAS
pathway.
 More specifically, it blocks the subtypes MEK 1 and MEK 2.
Jane PA, J Clin Oncol 2012; 30: Abstr 7503

64. SELECT 1 TRIAL
Selumetinib 75
mg bd continuous
plus docetaxel 75
mg/m2 on day 1 of
every 21 day cycle
v/s
Placebo plus
docetaxel 75
mg/m2 on day 1
of every 21 day
cycle
This study will assess the pharmacokinetics, safety, patient related outcomes
And tolerability profile of selumetinib/docetaxel compared toplacebo/docetaxel
INCLUSION CRITERIA
Male or female, aged 18 years or older
Histologically confirmed NSCLC stage III-IV
KRAS positive tumour sample
Failure of 1st line chemotherapy

65. BRAF MUTATION
Found in 2% of lung cancer
 Poor prognostic indicator

66. HER 2
 Activated by homo or hetero dimerisation.
 Amplification of HER2 is found in 2-4% of NSCLC patients.
 More prevalent in adenocarcinoma
 Negative prognostic marker
 Neratinib, a pan HER inhibitor has shown clinical activity in phase I trials

67. C-MET
 MET amplification occurs in 2-5% of NSCLC
patients.
 Potential mechanism of resistance in EGFR
inhibition.
 Ornatuzumab is a monoclonal antibody
against HGFR. In a phase II study,
ornatuzumab with erlotinib in comparision
to erlotinib with placebo as a means of 2nd or
3rd line therapy did not show any
improvement in survival.
 Tivatinib has high affinity for inactive kinase
domain of MET.
 The randomised, double blind MARQUEE
study investigated the addition of tivatinib
to erlotinib in NSCLC patients. After a
planned interim analysis, the trial was
stopped due to futility.

68. RET
 RET amplification occurs in 1.2% of
pateients with adenocarcinoma histology,
younger patients, light or never smokers
and poorly differentiated tumours.
 Alectinib is potent inhibitor of ALK and
shown anti tumour activity against RET.
 Other TKIs such as vandetanib,
cabozantinib, ponatinib have shown
activity against RET

69. FGFR1
 FGFR1 amplification occurs in 20%
squamous cell ca of lung and 5% of
adenocarcinoma.
 More common in male smokers
 Associated with poor outcome
 Clinical trials with FGFR inhibitors
are ongoing.

70. DDR2
 Discoidin domain receptor 2
 DDR2 mutation is found in 4% of NSCLC patients
 Multikinase TKI dasatinib has shown clinical activity against DDR2.

73. TOXICITY
Median onset of cutaneous toxicity is after 1-2 weeks of initiation of treatment
With maximum intensity at 2-3 weeks.
 Rash appears to be dose related and most patients experience transient mild to
moderate symptoms.
 Moderate to severe rash occurs in 10% patients.

76. CONCLUSION
 EGFR TKIs are the most extensively studied of targeted therapies in
NSCLC.
 Cells develop resistance to TKIs leading to development of 2nd
generation TKIs.
 In most studies, TKIs shows improvement in PFS without any benefit
in OS.
 In cost effective analysis, targeted therapies show very minimal gain in
live year and quality adjusted live year.
 Despite these, it gives an impetus to move from ready made therapy to
tailor made therapy.
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